Molecular analysis of aflR gene in Aspergillus flavus isolated from Iran

Aspergillus flavus produces the most potent carcinogens, aflatoxins, when it contaminates agricultural crops. aflR gene regulates aflatoxin-related genes and it has been identified in four species of A. flavus, A. parasiticus, A. sojae and A. oryzae. Contamination of agricultural commodities with aflatoxin is a grave risk to humans and animals’ health. Aflatoxin related genes are clustered in a 75 kb region of genome in A. flavus. Investigations obviously demonstrated that aflatoxin biosynthesis needs the aflR gene product and an entirely functional aflatoxin biosynthetic cluster. The purpose of the current study was to investigate the presence of the aflR gene in A. flavus. Material and methods. Forty-two A. flavus isolates including 10 references, 25 clinical and 7 environmental isolates were analyzed in this study. The isolates were identified by morphology. To characterize morphologically, the conidial arrangement, philiades, vesicles and conidiophores were observed microscopically. Using PCR, the aflR gene was amplified with primers aflR1 and aflR2. PCR were carried out to amplify an 800 bp DNA fragment of aflR gene. Some amplicons were sequenced. The sequences were searched in NCBI database and analyzed with MEGA5 software. Results and discussion. Out of 42 A. flavus isolates, an 800 bp band was amplified for 35 isolates. No band was observed for seven isolates including 4 clinical and 3 environmental isolates. Data analysis demonstrated that 100% of reference strains and 84% of clinical strains produced the expected fragment while it was only 57.14% for environmental isolates. The sequences had 100% identity with A. flavus aflR gene which was deposited in the NCBI database. Conclusion. In conclusion, molecular analysis of the aflR gene showed that this gene was not amplified from some strains of A.flavus; therefore, perhaps it lacks the gene or it is greatly abnormal. Additional researches are needed to verify whether the strains with lack of aflR gene have a loss of function in production of aflatoxin or other mechanisms of regulation exist

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IL-6 contributes to metastatic switch via the differentiation of monocytic-dendritic progenitors into prometastatic immune cells

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2021-002856 ◽

2021 ◽

Vol 9 (6) ◽

pp. e002856

Author(s):

Ksenia Magidey-Klein ◽

Tim J Cooper ◽

Ksenya Kveler ◽

Rachelly Normand ◽

Tongwu Zhang ◽

...

Keyword(s):

Lung Cancer ◽

Data Analysis ◽

Conditioned Medium ◽

Mononuclear Cells ◽

Fluorescent Protein ◽

Metastatic Potential ◽

Loss Of Function ◽

Tumor Spread ◽

Hematopoietic Stem ◽

Proteomic Screen

BackgroundMetastasis is the major cause of death in patients with cancer. Myeloid skewing of hematopoietic cells is a prominent promoter of metastasis. However, the reservoir of these cells in the bone marrow (BM) compartment and their differentiation pattern from hematopoietic stem and progenitor cells (HSPCs) have not been explored.MethodsWe used a unique model system consisting of tumor cell clones with low metastatic potential or high metastatic potential (met-low and met-high, respectively) to investigate the fate of HSPC differentiation using murine melanoma and breast carcinoma. Single-cell RNA sequencing (scRNA-seq) analysis was performed on HSPC obtained from the BM of met-low and met-high tumors. A proteomic screen of tumor-conditioned medium integrated with the scRNA-seq data analysis was performed to analyze the potential cross talk between cancer cells and HSPCs. Adoptive transfer of tumor-educated HSPC subsets obtained from green fluorescent protein (GFP)+ tagged mice was then carried out to identify the contribution of committed HSPCs to tumor spread. Peripheral mononuclear cells obtained from patients with breast and lung cancer were analyzed for HSPC subsets.ResultsMice bearing met-high tumors exhibited a significant increase in the percentage of HSPCs in the BM in comparison with tumor-free mice or mice bearing met-low tumors. ScRNA-seq analysis of these HSPCs revealed that met-high tumors enriched the monocyte-dendritic progenitors (MDPs) but not granulocyte-monocyte progenitors (GMPs). A proteomic screen of tumor- conditioned medium integrated with the scRNA-seq data analysis revealed that the interleukin 6 (IL-6)–IL-6 receptor axis is highly active in HSPC-derived MDP cells. Consequently, loss of function and gain of function of IL-6 in tumor cells resulted in decreased and increased metastasis and corresponding MDP levels, respectively. Importantly, IL-6-educated MDPs induce metastasis within mice bearing met-low tumors—through further differentiation into immunosuppressive macrophages and not dendritic cells. Consistently, MDP but not GMP levels in peripheral blood of breast and lung cancer patients are correlated with tumor aggressiveness.ConclusionsOur study reveals a new role for tumor-derived IL-6 in hijacking the HSPC differentiation program toward prometastatic MDPs that functionally differentiate into immunosuppressive monocytes to support the metastatic switch.

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Carbon catabolite repression gene creA regulates morphology, aflatoxin biosynthesis and virulence in Aspergillus flavus

Fungal Genetics and Biology ◽

10.1016/j.fgb.2018.04.008 ◽

2018 ◽

Vol 115 ◽

pp. 41-51 ◽

Cited By ~ 19

Author(s):

Opemipo Esther Fasoyin ◽

Bin Wang ◽

Mengguang Qiu ◽

Xiaoyun Han ◽

Kuang-Ren Chung ◽

...

Keyword(s):

Aspergillus Flavus ◽

Catabolite Repression ◽

Carbon Catabolite Repression ◽

Aflatoxin Biosynthesis

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Magnitude of Voriconazole Resistance in Clinical and Environmental Isolates of Aspergillus flavus and Investigation into the Role of Multidrug Efflux Pumps

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01022-18 ◽

2018 ◽

Vol 62 (11) ◽

Cited By ~ 8

Author(s):

Raees A. Paul ◽

Shivaprakash M. Rudramurthy ◽

Manpreet Dhaliwal ◽

Pankaj Singh ◽

Anup K. Ghosh ◽

...

Keyword(s):

Aspergillus Flavus ◽

Efflux Pump ◽

Efflux Pumps ◽

Azole Resistance ◽

Wild Type ◽

Multidrug Efflux ◽

Environmental Isolates ◽

Content Type ◽

Multidrug Efflux Pumps

ABSTRACT The magnitude of azole resistance in Aspergillus flavus and its underlying mechanism is obscure. We evaluated the frequency of azole resistance in a collection of clinical (n = 121) and environmental isolates (n = 68) of A. flavus by the broth microdilution method. Six (5%) clinical isolates displayed voriconazole MIC greater than the epidemiological cutoff value. Two of these isolates with non-wild-type MIC were isolated from same patient and were genetically distinct, which was confirmed by amplified fragment length polymorphism analysis. Mutations associated with azole resistance were not present in the lanosterol 14-α demethylase coding genes (cyp51A, cyp51B, and cyp51C). Basal and voriconazole-induced expression of cyp51A homologs and various efflux pump genes was analyzed in three each of non-wild-type and wild-type isolates. All of the efflux pump genes screened showed low basal expression irrespective of the azole susceptibility of the isolate. However, the non-wild-type isolates demonstrated heterogeneous overexpression of many efflux pumps and the target enzyme coding genes in response to induction with voriconazole (1 μg/ml). The most distinctive observation was approximately 8- to 9-fold voriconazole-induced overexpression of an ortholog of the Candida albicans ATP binding cassette (ABC) multidrug efflux transporter, Cdr1, in two non-wild-type isolates compared to those in the reference strain A. flavus ATCC 204304 and other wild-type strains. Although the dominant marker of azole resistance in A. flavus is still elusive, the current study proposes the possible role of multidrug efflux pumps, especially that of Cdr1B overexpression, in contributing azole resistance in A. flavus.

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Vibrio gazogenes inhibits aflatoxin production through downregulation of aflatoxin biosynthetic genes in Aspergillus flavus

PhytoFrontiers™ ◽

10.1094/phytofr-09-21-0067-r ◽

2022 ◽

Author(s):

Shyam L. Kandel ◽

Rubaiya Jesmin ◽

Brian M. Mack ◽

Rajtilak Majumdar ◽

Matthew K. Gilbert ◽

...

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolite ◽

Aspergillus Flavus ◽

Fungal Biomass ◽

Expression Profiles ◽

Health Hazard ◽

Gene Clusters ◽

Aflatoxin Contamination ◽

Aflatoxin Biosynthesis ◽

Control Samples

Aspergillus flavus is an opportunistic pathogen of oilseed crops such as maize, peanut, cottonseed, and tree nuts and produces carcinogenic secondary metabolites known as aflatoxins during seed colonization. Aflatoxin contamination not only reduces the value of the produce but also is a health hazard to humans and animals. Previously, we observed inhibition of A. flavus aflatoxin biosynthesis upon exposure to the marine bacterium, Vibrio gazogenes (Vg). In this study, we used RNA sequencing to examine the transcriptional profiles of A. flavus treated with both live and heat-inactivated dead Vg and control samples. Fungal biomass, total accumulated aflatoxins, and expression profiles of genes constituting secondary metabolite biosynthetic gene clusters were determined at 24, 30, and 40 h after treatment. Statistically significant reductions in total aflatoxins were detected in Vg-treated samples as compared to control samples at 40 h. But no statistical difference in fungal biomass was observed upon these treatments. The Vg treatments were most effective on aflatoxin biosynthesis as was reflected in significant downregulation of majority of the genes in the aflatoxin gene cluster including the aflatoxin pathway regulator gene, aflR. Along with aflatoxin genes, we also observed significant downregulation in some other secondary metabolite gene clusters including cyclopiazonic acid and aflavarin, suggesting that the treatment may inhibit other secondary metabolites as well. Finally, a weighted gene correlation network analysis identified an upregulation of ten genes that were most strongly associated with Vg-dependent aflatoxin inhibition and provide a novel start-point in understanding the mechanisms that result in this phenomenon.

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Strains Differentiation of Microsporum canis by RAPD Analysis Using (GACA)4 and (ACA)5 Primers

Polish Journal of Microbiology ◽

10.33073/pjm-2011-020 ◽

2011 ◽

Vol 60 (2) ◽

pp. 145-148 ◽

Cited By ~ 9

Author(s):

ANITA DOBROWOLSKA ◽

JOANNA DĘBSKA ◽

MAGDALENA KOZŁOWSKA ◽

PAWEŁ STĄCZEK

Keyword(s):

Molecular Analysis ◽

Tinea Capitis ◽

Rapd Analysis ◽

Tinea Corporis ◽

Microsporum Canis ◽

Fungal Strains ◽

Central Poland ◽

Molecular Studies ◽

Genomic Dnas ◽

Reference Strains

Molecular analysis of dermatophytes (based on PCR fingerprinting) revealed high clonal differentiation between the genus and species. Microsporum canis (zoophilic dermatophyte, belonging to genus Microsporum), responsible for most cases of tinea capitis in children, tinea corporis in adults and dermatophytoses in cats, is very unique in comparison with other dermatophytes. Results of most molecular studies show that there is no clonal differentiation within M. canis as distinct from other species. The aim of this study was application of (GACA)4 repetitive primer and (ACA)5 primer for typing of M. canis strains isolated from human and animals in Central Poland. Fungal strains: 32 clinical isolates of M. canis, originated from patients from Central Poland; 11 strains isolated from infected cats (6) and dogs (7), reference strains of M. canis (CBS 113480), T rubrum (CBS 120358), T mentagrophytes (CBS 120357) and E. floccosum (CBS 970.95). The genomic DNAs of the strains were used as a template in RAPD reaction. No differentiation was observed for the analyzed M. canis strains using (GACA)4 and (ACA)5 typing.

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Inhibition of Aflatoxin Biosynthesis by Organophosphorus Compounds

Journal of Food Protection ◽

10.4315/0362-028x-43.5.381 ◽

1980 ◽

Vol 43 (5) ◽

pp. 381-384 ◽

Cited By ~ 23

Author(s):

M. F. DUTTON ◽

M. S. ANDERSON

Keyword(s):

Aspergillus Flavus ◽

Organophosphorus Compound ◽

Organophosphorus Compounds ◽

Culture Fluid ◽

Aflatoxin Production ◽

The Other ◽

Aflatoxin Biosynthesis ◽

Inhibition Effect ◽

Type Activity ◽

Acetate Derivative

The effect of a range of organophosphorus and various other compounds on production of aflatoxin by Aspergillus flavus was investigated. Five organophosphorus compounds - Chlormephos, Ciodrin, Naled, Phosdrin and Trichlorphon- at concentrations of 20 and 100 μg/ml of culture fluid were found to have activity similar to Dichlorvos, in that they lowered the level of aflatoxin produced and caused formation of several anthraquinone pigments. Two of these pigments have not previously been described, one was named Versicol and a suggested structure is presented, whilst the other compound was shown to be its acetate derivative. A rationale is suggested for the required elements of structure, which are necessary for an organophosphorus compound to have Dichlorvos-type activity. Two unrelated compounds, ammonium nitrate and Tridecanone were also found to elicit Dichlorvos-type activity. It is likely that tridecanone or its breakdown products competitively inhibit enzymes involved in aflatoxin biosynthesis. It is possible that this inhibition effect explains the lowering of aflatoxin production in lipid-rich commodities infected by A. flavus.

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